Microbial Functional Genomics and Synthetic Biology

与这类微生物合作的遗传限制，包括其二倍体性质、不完整的性周期和非典型密码子的使用，阻碍了对真菌生物学和耐药性的遗传基础的研究。基于CRISPR-CAS的技术现在已经彻底改变了我们在这些微生物中进行有针对性的基因组编辑的能力。这项CRC研究计划将开发基于CRISPR的新型生物技术，以建立大规模基因文库，用于流行的真菌病原体白色念珠菌的功能分析。像其他真菌感染一样，白色念珠菌的治疗是出了名的具有挑战性，并代表着一个日益严重的公共卫生问题。本研究将开发一种基于CRISPR干扰(CRISPRi)的基因抑制系统，用于白念珠菌耐药的比较功能基因组学分析，以确定重要的耐药基因调控因素，并进一步将CRISPRi技术扩展到全基因组功能基因组抗真菌药物敏感性筛选平台。此外，这项研究计划将开发和优化两个新的平台：基于CRISPR激活(CRISPRa)的基因过表达系统，以及针对真菌物种不同遗传应用的有针对性的CRISPR碱基编辑系统。总而言之，这项CRC研究计划将把基于CRISPR的尖端功能基因组学应用于当前和新出现的真菌病原体，以破译介导耐药性的遗传机制。

Studying the genetic underpinnings of fungal biology and drug resistance has been hindered by genetic limitations associated working with this group of microbes, including its diploid nature, incomplete sexual cycle and atypical codon usage. CRISPR-Cas-based technologies have now revolutionized our ability to perform targeted genome editing in these microorganisms. This CRC research program will develop novel CRISPR-based biotechnologies to establish large-scale genetic libraries for functional analysis in the prevalent fungal pathogen, Candida albicans. Like other fungal infections, C. albicans is notoriously challenging to treat and represents a growing public health concern. This research will exploit a novel CRISPR interference (CRISPRi)-based genetic repression system for comparative functional genomic analysis of drug resistance in C. albicans in order to identify important genetic modulators of drug resistance, and further expand CRISPRi technology towards genome-wide functional genomic screening platform for antifungal drug sensitivity. Additionally, this research program will develop and optimize two novel platforms: a CRISPR activation (CRISPRa)-based genetic overexpression system, and a targeted CRISPR base editor system for diverse genetic applications in fungal species. Collectively, this CRC research program will apply leading edge CRISPR-based functional genomics to current and emerging fungal pathogens to decipher the genetic mechanisms mediating drug resistance.